Wang J, Boedeker J, Hobbs H H, White A L
Center for Human Nutrition, University of Texas Southwestern Medical Center at Dallas, Dallas, TX, USA.
J Lipid Res. 2001 Jan;42(1):60-9.
Efforts to develop an in vitro model system to analyze apolipoprotein [a] (apo[a]) gene transcription, mRNA translation, and protein secretion have been complicated by the limited tissue and species distribution of apo[a] and the presence of regulatory DNA sequences remote from the apo[a] transcription start site. In the current study we examined primary hepatocytes cultured from apo[a] transgenic mice as a model system for analyzing apo[a] biogenesis. Hepatocytes from mice transgenic for a yeast artificial chromosome (YAC) encoding the entire apo[a] gene in its own genomic context (YAC-apo[a] hepatocytes) were unable to maintain apo[a] expression beyond 48 h of culture. This suggests that the apo[a] promoter was not active in cultured YAC-apo[a] hepatocytes. In contrast, apo[a] expression was maintained for at least 7 days in hepatocytes cultured from mice transgenic for an apo[a] cDNA under control of the mouse transferrin promoter (transferrin-apo[a] hepatocytes). Pulse-chase experiments established that more than 80% of apo[a] synthesized by both transferrin-apo[a] and YAC-apo[a] hepatocytes was degraded prior to secretion, independently of the coexpression of human apoB.Thus, low secretion efficiency appears to be a general characteristic of human apo[a] proteins in mouse liver. Apo[a] secretion was increased somewhat (from 18% to 32%) in the presence of lipoprotein-containing serum. Transformed cell lines derived from transferrin apo[a] hepatocytes retained characteristics of apo[a] secretion similar to those observed in primary cells. Primary and transformed apo[a] transgenic hepatocytes may provide valuable additional models with which to study posttranslational mechanisms regulating apo[a] secretion. - Wang, J., J. Boedeker, H. H. Hobbs, and A. L. White. Determinants of human apolipoprotein [a] secretion from mouse hepatocyte cultures. J. Lipid Res. 2001. 42: 60;-69.
开发一种体外模型系统来分析载脂蛋白[a]（apo[a]）基因转录、mRNA翻译和蛋白质分泌的努力一直受到apo[a]有限的组织和物种分布以及远离apo[a]转录起始位点的调控DNA序列的影响。在当前研究中，我们检查了从apo[a]转基因小鼠培养的原代肝细胞，作为分析apo[a]生物合成的模型系统。来自转基因小鼠的肝细胞，该小鼠的酵母人工染色体（YAC）在其自身基因组背景下编码整个apo[a]基因（YAC-apo[a]肝细胞），在培养48小时后无法维持apo[a]表达。这表明apo[a]启动子在培养的YAC-apo[a]肝细胞中不活跃。相比之下，在由小鼠转铁蛋白启动子控制下的apo[a] cDNA转基因小鼠培养的肝细胞中，apo[a]表达至少维持7天（转铁蛋白-apo[a]肝细胞）。脉冲追踪实验表明，转铁蛋白-apo[a]和YAC-apo[a]肝细胞合成的apo[a]中，超过80%在分泌前被降解，与人类apoB的共表达无关。因此，低分泌效率似乎是小鼠肝脏中人类apo[a]蛋白的一个普遍特征。在含有脂蛋白的血清存在下，apo[a]分泌略有增加（从18%增加到32%）。从转铁蛋白apo[a]肝细胞衍生的转化细胞系保留了与原代细胞中观察到的apo[a]分泌特征相似的特征。原代和转化的apo[a]转基因肝细胞可能提供有价值的额外模型，用于研究调节apo[a]分泌的翻译后机制。 - 王，J.，J. 博德克，H. H. 霍布斯，和A. L. 怀特。小鼠肝细胞培养物中人类载脂蛋白[a]分泌的决定因素。《脂质研究杂志》。2001年。42: 60 - 69。
